Spinal Cord Regeneration in Zebra Fish

JOE FETCHO: My lab really focuses on how the brain and spinal cord produce behaviors. Most of our work deals with how movements are produced. The problem I want to tell you about is the problem of how you go about curing an injured spinal cord.
It's very difficult problem, because normally, it's not possible to see the response of the nerve cells to a particular treatment-- to watch how they respond and whether they start growing again, and whether they reconnect to the nerve cells that they would normally connect to.
And the problem is that most of the behaviors we look at involve activity in lots of nerve cells. But the tools we have to study them allow us to only look at electrical activity in one or a very few cells at a time.
We set out to solve that problem by using imaging approaches that allow us to literally watch the activity of nerve cells in an intact animal. And our strategy to do that was to use an animal that's transparent-- the larval zebrafish.
So if we look at a picture of a larval zebrafish, this is what they look like. They're about 4 millimeters long. And you can get the sense that the animal's fairly transparent. The head's obviously over here. The spinal cord is on the top, and then the notochord.
If we zoom up on the tail region of the fish, we can see the transparency. This is a movie of a fish. This is the spinal cord here, and the notochord there. And if I animate the movie, you'll see that we can actually see the blood cells flowing through the capillaries in the nervous system of the fish.
This transparency is what allows us to look at the structure of neurons in the animal and to evaluate the effect of treatments on restoring the ability of the nerve cells to regrow.
Now, our work was focused on a nerve cell up in the brain that has a process and axon that goes all the way down the spinal cord that's involved in producing an escape response that fish use to avoid predators. It was a good cell to use, because it's big and it has a very long axon that goes all the way from the brain down to the tail of the fish.
This is what one looks like in a transparent live larval fish. And in this case, we filled it with dye-- red dye. So the head of the fish is on this side, and the tail on the right side. So the head on the left, the tail on the right. And this is the cell body of the cell. And this is its axon running down the spinal cord toward the tail.
This is actually a 3D reconstruction of the cell. So if we have it rotate, we can see that we can see all aspects of the neuron in the live fish. The strategy we adopted, then, was to damage this cell's axon by severing the spinal cord about here-- about 2/3 of the way toward the head from the tail.
And if we do that and we look at this cell after that damage, it looks like this. Here's the cell body up in the head. Here's the axon. And we've severed it here. And the part beyond where we severed it-- near the tail-- is degenerating.
Now, this cell would never grow a new axon on its own. It would sit there like that and function would not be restored, because it would not be able to regenerate the part that died after the damage. And the strategy was to see if we could induce it to regrow by testing different drugs.
And the way we tested them was we squirted the drug on the cell body-- up in the brain. And if we just squirt that on the cell, which we can see in the live fish, and we come back the next day, a cell that wouldn't regenerate and was sitting here has now grown an axon that goes down much further in the spinal cord.
So we've induced the cell to grow through the lesion and down the spinal cord, which is really what a cure for spinal injury is going to take. The other advantage that we have is, because this is an intact animal, we can actually watch the growth process itself.
So we can collect images like this every hour for 36 hours, for example. So if we look at this site right here-- where it's been cut-- and just look at the cut end, which we see here is the cut end of the axon. And we take an image every hour for 36 hours, here's what we see.
That cut region sprouts prostheses. One of those prostheses makes it across the lesion sites. So the head end is to the left, and the tail to the right. If I run it through again, it sprouts prostheses. And this one makes it across the lesion site.
So within about 36 hours, it's sprouted and grown across the lesion site. And now if we come back and look at it in the view of the whole cell-- here's the cell body in the brain. Here's the site I just showed you where it regrew across the lesion here. And we see that not only did it go across, but it grew down the spinal cord.
So again, we have a very robust strategy for inducing regeneration that is easily evaluated, because we just watch it happen. The behavior that this fish produces is an escape behavior, and that escape behavior is shown here.
These fish are very fast. They move so quickly, you can't see them do this escape behavior with your eyes. So we have to film them with cameras that work in 1,000 frames a second that were originally developed for filming explosions.
This is an example of one of those films where there's a squirter up here that's going to squirt the fish in the head. This whole sequence is about 40 milliseconds long-- 40 thousandths of a second. It squirts the fish in the head, and the fish does this turn and swims away. It normally uses this response to avoid being eaten by predators.
The cell I showed you is critical for this response, and the fish can't do this response properly when that cell's been damaged. So if we look at a fish on the left here, which is a fish that's been lesioned where that cell's axon's been severed, and we squirt it in the tail-- we'll see that it doesn't get away very well.
It doesn't bend for a time. And then it does a very weak bend. And it doesn't swim away very well. Its escape response is hugely impaired.
If we then squirt the drug on the cell, get it to regrow, and take the same fish, now, on the right side and squirt it, we see that the response recovers. The fish does a very robust bend and swims away effectively.
So not only have we restored the ability of the cell to regrow, that cell has also reconnected in a way that appears to restore the normal function of that nerve cell.
I think it's very important to appreciate that we can learn a lot about human biology by studying non-human animals. We can learn a tremendous amount even from studying fish, as I have for a long time. Because we share with fish a lot of basic features of our biology, because we're all related by evolution.
And even though a fish may look very different, a lot of the organization of its nervous system and processes related to regeneration and normal functioning are very similar to us. I think, to some extent, society's losing sight of the relevance of all animals for human biology.

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Zebra fish are unique because their larvae are transparent and their nervous system can be observed through their skin.

Cornell neurobiologist Joe Fetcho uses zebra fish as a model organism for developing a basic set of principles on the organization and function of neurons, and for studying cell regeneration. He has developed a robust strategy for inducing cell growth and restoration of function, both necessary for curing spinal cord injury.

In September 2009, Fetcho received the NIH Director's Pioneer Award, which supports scientists of exceptional creativity who propose pioneering approaches twho propose pioneering approaches to major challenges in biomedical and behavioral research.